1. Field of the Invention
This invention relates to an apparatus and method for detecting the refractive index and flow rate of a fluid, and particularly to such apparatus and method for use as detectors in liquid chromatography.
2. The Prior Art
Liquid chromatography pertains to a particular variety of equipment and techniques for analyzing the components of an unknown sample of liquid material.
Liquid chromatography is a process wherein a sample having unknown components is forced to migrate through an elongated "column". The column contains a material held statically inside it, called a "stationary phase". The stationary phase is chosen for its ability to selectively retain the various potential components of the sample with which it comes in contact with differing degrees of tenacity. The sample is forced to migrate through the column by injecting it into a solvent upstream of the column and subsequently pumping the solvent and dissolved sample through the column.
When the dissolved sample is forced through the column, each of its components migrates through the column in a particular time related pattern, which pattern is a function of the degree of the tendency of the stationary phase to retain that component.
Some properties of the column effluent, following the pumping of the solvent and dissolved sample through the column, are affected by the concentration of sample in the emergent fluid. One of the properties which is typically so affected is the refractive index of the effluent. By detecting variations in such properties of the column effluent, and by plotting these variations against time, certain information can be derived as to the nature and amount of the components in the sample.
For example, for predetermined column conditions and flow rate, it may be known that a particular hypothetical component, if present in the sample, will reach a maximum concentration in the column effluent at a specific time following introduction of the sample to the column. This time is known as the "retention time" of the component. This phenomenon occurs because of the existence of a particular degree of retention of that component by the stationary phase. By measuring a property of the effluent known to be affected by the hypothetical component, and observing whether a maximum occurs at the "retention time" for that component, the presence of the component can be verified or negated.
Conditions of the effluent affect precise determination of some hypothetical components, such as effluent flow rate affecting the retention time of the component. Therefore the flow rate is desirably known even in systems inconvenient for flow rate measurement.
It is evident that in liquid chromatography there exists a necessity for accurately detecting and measuring, on a continuous basis, properties of the liquid emerging from the column, such as refractive index. It is further evident that a necessity exists for determining effluent conditions in liquid chromatography systems such as fluid flow rate, notwithstanding inconvenience of measurement.
Several types of apparatus exist for detecting changes in the index of refraction of the column effluent. One (called "deflective" type) involves passing the column effluent through an elongated flow cell having a triangular cross-section, the hypotenuse of which triangle forms an interface with that of a second triangular cross-sectional chamber. The second chamber contains a reference fluid having a known index of refraction. A mirror is placed parallel to one of the legs of the second chamber at a distance therefrom. A light beam is then directed through the two chambers, and across the interface, at which point it is bent, and transmitted on to the mirror at an angle of incidence dependent upon the difference between the indices of refraction of the column effluent and the reference fluid. The light beam is reflected from the mirror and returns back across the interface, being bent additionally as it crosses the interface to an angle even further removed from the angle at which the incoming light beam was incident on the interface for the first time. The degree of deflection of the light beam is measured, and is a function of the difference between the respective indices of refraction of the column effluent and the reference fluid.
Another type of refractive index detector employs two beams of light which originate from a common region of a tungsten filament lamp. The two parallel beams of light pass through a glass prism, and are partially transmitted through two glass-liquid interfaces. One interface is the boundary of the detection flow cell and the prism, and the other interface is the boundary of a reference cell with the prism. The two transmitted beams of light are then scattered from a finely ground stainless steel back plate, and a part of the scattered light from each of the beams is transmitted back through its respective cell, the glass prism and on to two halves of a photoconductive sensing cell.
The ratio of the amount of light transmitted through the two interfaces is a function of the refractive indices of the substances in the two cells. Thus, measurement of the transmitted light may be used to derive the refractive index of the substance in the detection cell, provided the refractive index of the material in the reference cell is known.
The deflection type of detector described above is less susceptible to changes in solvent composition than is the reflection type of detector. On the other hand, the deflection type of detector offers a smaller linear range than does the reflection type. Thus, prior art detectors have not fully combined the advantages of the reflection and deflection detectors.
Both of these principal refractive index detectors are adversely affected by changes in temperature of the liquid passing through their flow cells. This is because the refractive index of most liquids is dependent to some degree upon temperature. Therefore, measures must be taken to provide for temperature compensation of these devices. This technique adds to the complexity and expense of the detection instrument.
Various types of flow meters have been proposed for measuring flow rate of a fluid through a passage. Flow meter proposals typically require a member physically disposed in the fluid and linked to a measurement device. Oftentimes such a member is not conveniently combined with the system in question whose fluid flow rate is unknown.